The present invention relates to treated primed seeds having a longer shelf life than conventionally primed seeds, a process for obtaining such seeds and plants derived therefrom.
Primed seeds and methods for obtaining them are known in the art.
Primed seeds are generally capable of a faster germination over time and display better synchronization in germination, than non-primed seeds.
Conventional seed priming processes are i.a. disclosed by EP 309 511 B1 and EP 254 569 B1.
The present invention provides a method of treating primed seeds which results in a prolongation of shelf life compared to optionally dried back conventionally primed seeds of the same species having substantially the same moisture content (MC) characterised in that the primed seeds arc subjected to a water stress, a heat treatment or a combination thereof and are subsequentlyxe2x80x94where desiredxe2x80x94dried back to a desired MC.
The drying back of the seeds as abovementioned, involves a reduction in the seed MC. The drying back may be to any desired MC. In a preferred embodiment the drying back is to a MC of untreated seeds, i.e. dry seeds which have not been primed.
Hereinafter the terms MC and water content will be used interchangeably. The MC will be given in percentage terms, based on seed fresh weight(unless otherwise stated).
The water stress may be achieved by any manner known in the art which will result in the seeds having a lower MC.
In general a water stress will be obtained by reduction of the MC of conventionally primed seeds by 5% units or more, i.e. to 20% or less, if the conventionally primed seeds have an initial MC of 25%, or to 50% or less if the conventionally primed seeds have an initial MC of 55%. More specifically, the water stress will in general be achieved by reduction of the MC by 5% units up to 20% units, whereby, it is in general advantageous not to reduce the MC to a value below 15%
The water stress should be maintained for a longer period of time, in general 1 to 7 days, depending i.a. on the temperature, whereby the optimum conditions will depend on the seed species and can be determined by standard tests. It will be appreciated that this can be achieved by maintenance of the MC of the conventionally primed seeds constant at the desired reduced level (i.e. conveniently 5 to 20% units below the MC of conventionally primed seeds) or by drying the conventionally primed seeds slowly enough for them to remain sufficiently long under water stress.
Accordingly prolonged shelflife may be attained by incubation of the primed seeds at a water potential which induces a water stress, by slow MC reduction of primed seeds, by an initial quick MC reduction of primed seeds up to a MC where the seeds are still subject to water stress, followed by incubation or slow MC reduction of the thus obtained partly dried primed seeds or by a heat shock. Slow MC reduction can be achieved in a manner known per se, e.g. by drying under mild conditions or by bringing the primed seeds in contact with an osmoticum that is not toxic to the seeds and having a water potential below O MPa. The description of processes (a), (b) and (c) hereinafter, illustrates typical conditions for subjecting primed seeds to a water stress.
The water stress may be achieved by:
a) slowly drying primed seeds at a temperature of from 3 to 40xc2x0 C. for 3 to 7 days, or
b) reducing the MC of primed seeds under conventional drying conditions by 5 to 20% units and storing the thus dried seeds for 1 to 7 days in a container with minimal air and moisture exchange, at a temperature of from 3 to 40xc2x0 C. or
c) incubating primed seeds in an osmoticum for 1 to 7 days at a water potential chosen to reduce the MC of the primed seeds by 5 to 20% units.
The heat treatment may be achieved by subjecting primed seeds to a heat shock in the range of from 25 to 45xc2x0 C. for from about 1 to 5 hours.
The seeds produced according to the processes of the invention, have greater desiccation tolerant embryos than conventionally primed seeds, in that the former survive longer storage periods under ambient storage conditions as hereinafter defined.
By a seed comprising a desiccation tolerant embryo is meant a seed in which a reduction the MC of the seed to a value typical for dehydrated seeds e.g. of about 5 to 7%, does not substantially adversely affect the seeds viability, the viability being measured in terms of ability to germinate when placed under suitable growing conditions or after an appropriate standard test, e.g. a controlled deterioration test (see hereinafter) either before or after a prolonged storage period at ambient storage conditions.
The embryo of the seed is taken to mean structures which are necessary for the development of the seed, such as the cotyledon, axis and non-emerged radicle tip and which collectively or in part are able to acquire a desiccation tolerance.
Primed seeds can be stored for a number of weeks at about 5xc2x0 C. but are unsuitable for storing for extended periods of time under ambient storage conditions.
By the term primed seed is meant (insofar as not otherwise stated), that the seed has been subjected to conventional priming techniques as hereinafter discussed, it has a MC in the range of from 20 to 55% (depending on species) and has a desiccation tolerance typical of conventionally primed seed. It will be appreciated that all non-primed seed is desiccation tolerant i.e can survive drying, the extent of desiccation tolerance being species dependent. On priming according to conventional processes, the seed becomes less desiccation tolerant, the loss in desiccation tolerance increasing with the increase in period of priming, until a point at which the seed can no longer be said to be desiccation tolerant, this complete loss of desiccation tolerance occurring at the point of germination of the seed. The processes of the present invention, as hereinafter described, are applied to non-germinated seed which has been primed according to conventional priming processes. Non-germinated seed is defined herein as seed in which the radicle and/or hypocotyl has not protruded or emerged from the seed coat or pericarp. The radicle and/or hypocotyl may have caused the seed coat to split or crack, however it will not have protruded through the split or crack. The endosperm surrounding the embryo, may be visible through the split or crack. Non-germinated conventionally primed seed to which the proceeses of the present invention have been applied, will hereinafter be referred to as as treated seed(s). Non-germinated conventionally primed seed(s) to which the processes of the present invention have not been applied, will hereinafter be referred to as conventionally primed seed(s). Commercially acceptable seed(s) which have not been primed, shall be referred to as untreated seed(s).
It is also possible to determine the stage of the germination process by physical parameters, e.g. size, volume or density. In this way, a selection can be made of seeds to be treated according to the invention.
Thus as is demonstrated below, treated seed has a longer shelf life than conventionally primed seed of the same species and MC. The longer shelf life can be demonstrated by measuring % germination under the same or similar conditions e.g standard growing conditions (as below defined), after either a controlled deterioration test, or storage under ambient conditions; the treated seed having higher % germination of normal plants compared to conventionally primed seed which have been subjected to the same controlled deterioration test or storage conditions.
By standard growing conditions is meant a temperature in the range of about 15 to 20xc2x0 C. in the presence of air and water.
The term shelf life as used herein may be expressed in terms of viability (i.e. in terms of ability to germinate and give rise to normal plants after storage under ambient storage conditions, for example after subjection to a controlled deterioration (CD) test (Tarquis A. M. and Bradford K. J., J. Exptal. Bot. Vol. 43, 1982, No. 248,pp. 307-317). The viability of seeds subjected to the CD-test may be determined in laboratory tests according to International Rules (ISTA, 1976). Differences between CD test results generally correlate with differences in shelf storage life after storage under ambient storage conditions.
Typical examples of conventional priming methods include treatment of seeds with an osmoticum (as disclosed i.a. by Heydecker, and modifications thereof, such as for example the Drum Priming Method), treatment with water in a solid matrix (as disclosed i.a. in EP 309 551 B1) etc.
By the term xe2x80x9cambient storage conditionsxe2x80x9d is meant storage at ambient temperature and relative humidity (RH).
The term xe2x80x9cambient temperaturexe2x80x9d as used herein, refers to a temperature from about 3xc2x0 C. to about 25xc2x0 C. By the term xe2x80x9cambient RHxe2x80x9d is meant a RH in the range of from about 20% to about 90%.
Application of the processes (a), (b) or (c) of the invention as hereinbelow described, involves a reduction in seed MC.
Process (a) of the invention involves a slow rate of water loss of primed seeds(hereinafter referred to as slow drying). Thus, the primed seeds are subjected to an incubation phase in which the rate of water loss is maintained within the range of from about 0.1% and 1.0% of dry weight of the seed hourxe2x88x921, preferably within about 0.2% to about 0.4% hxe2x88x921. Slow drying can be performed in a drum (drum priming), oxygenated gas or oxygen being actively supplied or air may enter the system (passive conditions) simply, by mixing. The rate of water loss is determined by weighing seeds after different periods of incubation and plotting seed weight over time. It will be appreciated that the rate of moisture loss required to induce long shelf life will vary, from species to species, within the above defined range. Seeds held under such conditions will have a final MC of between about 5% to about 20%, generally from about 5% to about 15%, lower than that of the primed seeds.
In slow drying, the seeds can be incubated at any temperature between 3xc2x0 C. up to about 40xc2x0 C. Preferably, seeds are incubated at temperatures from about 20xc2x0 C. to about 35xc2x0 C. The incubation period lasts for about 24 hours up to about a week or more depending on incubation temperature. Thus e.g. the incubation period may be from about 24 hours up to about 3 days or more, at a temperature of 20xc2x0 C., depending on seed type, or it may be for up to a week or more at a lower temperature, such as 8xc2x0 C., depending on seed type. Lower temperatures can be employed, for example, to minimise the risk of infestation with pathogens.
According to process (b), hereinafter referred to as moist storage, primed seeds are incubated at a seed MC which is lower than the seed MC of primed seeds. Thus the MC of primed seeds is reduced (by drving under conventional drying conditions e.g. xe2x80x9cfast drying conditionsxe2x80x9d), by between 3 to 20% units, preferably between 5% and 15% units, over a period of less than 24 h. such as 8 h or less. The minimum value MC to which the seeds are dried is about 15%. Thereafter the so dried seeds are subjected to a water stress by incubation in a container with minimal air and moisture exchange, the temperature and length of incubation being a, for slow drying above.
The term conventional drying conditions as used in connection with process (b) hereinabove (and where used hereinbelow) refer to conventional drying conditions e.g. drying back by means of a high speed air flow at ambient temperature as known in the art and generally applied for the drying of conventionally primed seeds.
A further method of subjecting primed seeds to a water stress, the incubation process according to process (c) of the present invention, comprises a PEG (or other suitable osmoticum solution) treatment, which involves a reduction in the seed MC of seeds which have been subjected to a priming step by incubation in solutions, the water potential of which is less than O MPa. During incubation seed water content is slowly reduced by between 3 to 20% units, as described for process (b), by keeping the osmotic potential of the solution at a specified value, within the range of from about xe2x88x920.5 to about xe2x88x924.0 MPa. In this state the seeds experience a mild water stress due to a lack of availability of free water. Incubation preferably takes place in a water column (liquid incubation), preferably under aerated conditions. It can also take place by placing the primed seeds on filter paper saturated in an osmoticum solution.
Suitable incubation conditions (length, temperature) for process (b) and (c) include i.a. the conditions as described for process (a).
The incubation according to process (c) will typically be carried out in an osmoticum having a water potential that is low enough to withdraw water from the primed seeds. Any suitable osmoticum which does not harm the seeds, can be used e.g. polyethylene glycol (PEG) solutions such as PEG 8000 (British Petroleum). Typically, the seeds are contacted with a solution such as PEG 8000, mannitol, or a salt solution such as NaCl or the like. The osmotic potential should be such, that the seed MC is held at a sufficiently low level so that the induction of desiccation tolerance occurs.
Plant growth regulators may be added to the osmoticum solution at a concentration of between about 10xe2x88x922 M and 10xe2x88x928 M in the first incubation. Suitable regulators include giberellins, abscisic acid (ABA), and auxins such as indole butyric acid (IBA).
In incubation in a water column, the amount of seed per unit volume solution, can be from 1-200 g seeds 1xe2x88x921. Preferably, the seeds are present at about 25 g seeds 1xe2x88x921. Generally, the incubation may be for several days, extending to weeks or longer. After incubation, the seeds are washed in water.
With incubation on filter paper, the paper is moistened with a suitable osmoticum (as above described). Generally, seeds imbibed and primed to a MC of between about 25% and 55%, can be laid onto the moistened filter paper in a closed system, having a high RH eg 100%, the temperature and contact time being as above described).
Where desired the MC of primed seeds can be reduced by e.g. about 10% units by fast drying, before incubation in an osmoticum as above described. Such a step is not however essential.
In the heat treatment, the primed seeds are subjected to a heat shock in the range of about 25 to 45xc2x0 C., preferably in the range of 35 to 40xc2x0 C., for periods of time in the range of from about 1 to about 5 h. Preferably the MC of seed which is subjected to a heat shock is in the range of from 0 to 20% units lower than the seed MC after priming. Generally it is advantageous that the MC of the seeds which are subjected to a heat shock, is not lower than 15%. The heat shock may be applied by any known suitable method. Thus suitably the seeds may be placed in a container which is then placed in an incubator.
It will be appreciated that the desired result (primed seeds having a long shelf life), may also be attained by a combination of water-stress and heat-stress i.e. a combination of heat shock with either process (a), (b) or (c).
The optimum process and optimum conditions for the above mentioned process (a) to (c) and the heat shock for a given seed species, may be established by determining the shelf life potential and germination rate (t50) in as ma known per se.
Treatment of primed seeds according to any of the processes (a) to (c), the heat shock or a combination of such processes as above described, and subsequentlyxe2x80x94where desiredxe2x80x94drying the seeds back to a desired MC, gives rise to seeds having a longer shelf life than conventionally primed seeds of the same species having substantially the same MC.
The invention therefore provides seeds obtainable by process (a), (b), (c), the heat shock or a combination of such processes as above described, the seeds subsequentlyxe2x80x94where desiredxe2x80x94being drying back to a desired MC.
The present invention also provides treated primed seeds in wet or dry form, having a shelf life, when stored under ambient storage conditions, which is substantially longer than that of conventionally primed seeds of the same species having substantially the same MC and of which the MC has optionally been reduced under conventional drying conditions.
The seeds of the invention have a MC within the range of that typical for dry i.e untreated seeds, up to a MC at which metabolic processes other than germinative metabolic processes continue.
Typical commercial forms of the seeds of the invention include seeds having a MC in the range of from more than 15% to about 55%, (hereinafter referred to as wet seeds of the invention) and seeds that have been dried back to roughly the MC of dry seeds, i.e. having a MC in the range of from about 2% to about 15% of seeds (hereinafter dry seeds of the inventions).
Dry seeds of the invention are obtained by drying back seeds obtained according to any one processes (a), (b), (c), a heat shock, a combination of either (a), (b) or (c) with a heat shock, according to the invention, to a final MC of the order of that of non-germinated, non-primed seeds (i.e. the untreated seeds) using conventional i.e. fast drying conditions. Under conventional drying conditions, seeds can be dried back at a temperature lying within the range of from 10xc2x0 C. to 50xc2x0 C. generally from 20xc2x0 C. to about 35xc2x0 C., at a relative humidity within the range of from 30% to 90%, generally from 30% to about 50%, in still air or in flowing air at speeds typical for drying back seeds. For example, the airflow speed may be at any speed up to 2 m sxe2x88x921 or faster. The period of time may be for any suitable time interval up to 24 hours, depending on drying conditions employed. Suitable conventional drying conditions include, for example, temperature of 20xc2x0 C., a relative humidity of 40% in air flowing at a speed of 2 m sxe2x88x921 over 16 hours.
The dry seeds of the invention are useful, in that their germination rate is substantially shorter than that of untreated seeds of the same species. The germination rate is typically expressed in terms of t50, i.e. the time by which 50% seeds of a seed sample germinate.
The wet seeds of the invention are useful, i.a. in that they can be dried back to dry seeds of the invention in conventional manner.
Thexe2x80x94dry and wetxe2x80x94seeds of the invention have furthermore the advantage that they have a substantially longer shelf life than conventionally primed seeds of the same species having substantially the same MC.
The invention accordingly provides non-germinated seeds having a MC in the range of from 2 to 55% characterized in that said seeds when having a MC of untreated seeds, or after having been dried back to such a MC under conventional drying conditions, have a t50 which is substantially shorter than that of untreated seeds of the same species. Suitably the t50 is 60% or less than that of untreated seeds of the same species. Preferably the t50 is 50% or less, more preferably less than 40%. The MC of untreated seeds is generally in the range of from 2 to 15%. The t50 of the dry seed of the invention is generally substantially the same as the t50 of conventionally primed seed of the same species having substantially the same MC.
The t50 may be determined in a conventional manner e.g. according to the method of Orchard T. G. ( 19771 ) Seed Sci. and Technol. Vol 5, pp. 61-69. Generally such a determination is carried out at a temperature in the range of from about 15 to 20xc2x0 C. on e.g water saturated filter paper.
The invention further provides treated primed seeds in wet or dry form having a shelf life when stored at ambient storage conditions which is substantially longer than that of conventionally primed seeds in either wet form or after having been subjected to conventional drying conditions.
The term shelf life as used herein refers to the time period (term) that seeds can be stored under ambient conditions without substantially losing their ability to germinate.
The ability of seeds of the invention to germinate is accordingly substantially unaffected after storage over a period of time and under conditions which adversely affect the germination ability (expressible in % normal plants germinating) of conventionally primed seeds.
For convenience, it is accepted that seeds have not substantially lost their ability to germinate after storage, if the % of germinating plants has not been reduced by more than 20% units, preferably less than 15% units, more preferably less than 10% units, after storage.
Thus, by the statement xe2x80x9cseeds of the invention have a shelf life of at least 35% longer than conventionally primed seedsxe2x80x9d is meant that it takes at least 35% storage time units longer for seeds of the invention to substantially lose their germination ability (% normal plant germinating) than conventionally primed seeds of the same species stored under the same conditions.
Dry seeds of the invention have a shelf life which is substantially longer than that of conventionally primed seeds of the same species of the same species having substantially the same MC. Suitably the shelf life is at least 35%, more specifically at least 50% longer than that of conventionally primed seeds of the same species when said conventionally primed seeds are dried back to a MC of untreated seeds under fast drying conditions as typically employed for the drying of conventionally primed seeds. Under optimum incubation/heat treatment conditions, the shelf life may be extended by 150% and more. Typically the shelf life will be extended by 50 to 120%, and even after incubation/heat treatment under less optimized conditions, by 50 to 100%. In absolute terms, the shelf life of dry seeds of the invention will easily exceed 8 months, more specifically 12 months, when stored under conditions typical for untreated seeds, and extend to 24 months or longer. The dry seeds of the invention have preferably a MC in the range of from 5% to 8%. The shelf life of the seeds of the invention, will not be longer than that of untreated seeds of the same species.
Suitable storage conditions for the dry seeds of the invention are ambient storage conditions, for untreated seeds e.g at a relative humidity generally of from about 20% to 90%, preferably from about 30% to 60% and a temperature of from about 3xc2x0 C. to 25xc2x0 C., depending on the seed type.
Appropriate shelf storage conditions for wet seeds of the invention may comprise storage in a container with minimal air and moisture exchange at a temperature of from about 3xc2x0 C. to 10xc2x0 C., depending on seed type. Under such storage conditions the seeds have a shelf life of from about 4 to 6 weeks.
The seeds of the invention may be any desired seed species to which a conventional priming process can be applied. Examples of suitable seed types include tomatoes, peppers, melons, water melons, cucumbers, Brassicas, leeks, carrots, onions, squashes, gherkins, endives, Impatiens. Verbenas, Primulas, Pelargoniums, Viola, Chigoriums and Cyclamen. Specific examples of Brassicas are cabbage, broccoli, cauliflower and Brussel sprouts.
Also encompassed within the ambit of the present invention are plants grown from seed as herein described.
There are a number of methods of priming seeds known in the art. These are briefly reviewed hereinafter:
Non-primed i.e. untreated seeds, depending on species, may be soaked up to a few hours in an aqueous solution in e.g. a water column, in a pre-priming treatment. Such a pre-treatment known in the art, helps prevent the seeds from sticking together during priming, and/or readies the seeds for priming.
Non-primed seeds or seeds which have undergone the aforementioned pre-priming treatment, are placed under conditions e.g. time, temperature, water uptake by seed, which enable the seed to imbibe water to a level at which pre-germinative metabolic processes commence and continue but at which germination(as above defined) is not possible.
Imbibition may be carried out according to any known imbibition process. Thus for example (non-germinated) seeds to be imbibed may be placed in a drum or a water column, with or without aeration, at a water potential of 0 MPa (if the seeds are placed in water), or between about 0 MPa to about xe2x88x921.5 MPa if the seeds are placed in an osmoticum solution. Depending on the choice of priming technique, the amount of water imbibed is defined by the osmotic potential of the priming solution (in aqueous liquid priming techniques such as water columns) and the amount of water added to the system (for eg drum priming techniques).
The seeds imbibe added water until their MC typically rises to between about 25% to about 55%, preferably to about 30% to about 50%, depending on seed type.
MC of seeds is calculated using the formula:             Wi      -      Wa        Wi    xc3x97  100
where
Wi=weight initial
Wa=weight after oven drving seeds at 103xc2x0 C. for 16 hours or 130xc2x0 C. over 2 hours.
Imbibition takes place at any temperature conducive to the up take of water, generally, between about 5xc2x0 C. and about 30xc2x0 C., depending on species. When imbibition takes place in a water column, the degree of aeration should be sufficient to keep the seeds buoyed or in suspension. Imbibition can be for any suitable period up to about 24 hours, preferably from about 4 to about 10 hours depending on species. It may be a separate step before priming or may be an integral part of priming.
For proper priming the MC of the seeds is maintained at a relatively constant level, ie xc2x11 to 3% of the desired MC, typically i.e. between about 20% to about 55%, preferably between about 30% and 50%, of the seeds. Preferably, priming is carried out in a drum, for about 1 to about 21 days, preferably from about 2 days to 15 days, typically at a temperature in the range of about 5xc2x0 C. to about 30xc2x0 C., preferably from about 15xc2x0 C. to about 25xc2x0 C., depending on species.
The optimum seed MC and length of the priming step, depends on the particular seed type employed. These optimum values can be found using conventional procedures, for example, by setting different MCs for seeds, subjecting seeds to different incubation periods under certain controlled conditions eg temperature, RH and aeration.
Where it is desired to add a biological to the seed, the biological may be applied using techniques known in the art. Thus for example the biological may be added in any suitable form e.g an inoculum which may/may not be in the form of a suspension of micro-organisms in a suitable medium, dry fungal spores or freeze-dried or lyophilised bacteria. The biological may be added at any suitable stage of the process of the invention. Preferably it is added in the form of an inoculum at or near the beginning of the priming or alternatively in processes involving a treatment before priming, at or near the beginning of such pretreatment.
Suitable biologicals may be selected from the group comprising beneficial micro-organisms such as Bacillus. Pseudomonas, Trichoderma and Rhizobia. Particular examples of suitable micro-organisms include Pseudomonas fluorescence, Pseudomonas putida, Xanthomonas maltophilia, Bacillus spp. such as Bacillus subtilis, Bacillus thuringiensis, Bacillus cereus, Trichoderma viride, Trichoderma harzarium, Trichoderma koningii, Gliocladium virens, Fusarium oxysporum(non-pathogenic isolates) and the like. The biological may be specifically chosen where it is desired to treat a seed such that it is resistant to a particular plant pathogen e.g. Pseudomonas may be added to seed which is for sowing in soil known to be heavily infested by Pythium or Bacillus spp. may be added to seed to be sowed in seed which is prone to attack by Alternaria spp. eg carrot.
Generally the biological should be present in the range of from 103 to 109 colony forming units(cfu) seedxe2x88x921, depending on species of seed and biological. Thus e.g the cfu of Rhizobia on legumes such as alfalfa, should be about 103 seedxe2x88x921. However for most biologicals the cfu seedxe2x88x921, can be in the order of about 104 to about 107.
Where a seed coating is to be applied to the seed, it may be applied before, after or during the incubation period and either before or after any subsequent drying step. The coating may comprise any conventional material commonly used in seed coatings and may be added to the seed using conventional coating or pelleting techniques. The coating may comprise plant growth regulators such as gibberellins or auxins and/or any of the abovementioned micro-organisms such as Pseudomonas or Trichoderma and the like. Typically, the content of growth regulator will be in the range of from about 0.0001% to about 0.1% by weight of the coating material.
The coating may comprise any conventional material commonly used in the art for protecting or pelleting seed. Suitable materials include clays such as sub-bentonite and bentonite, vermiculite, along with additives such as perlite, pumice, metal stearates, polyethene, polystryrene, polyurethane, talcum powder, polypropene, polyvinyl chloride, starches, loams, sugars, arabic gums, organic polymers, celluloses, flours such as wood flours, quartz powders and the like.
Accordingly the present invention further provides seeds obtainable by any one of the abovementioned processes, the seeds being colonised with beneficial biologicals.
In a yet further embodiment the present invention provides seed obtainable by any one of the abovementioned processes, the seed being provided with a protective coating which optionally has added biological.
Then no follow examples which further illustrate the invention. It is to be understood that the examples are not to be viewed as limiting the scope of the invention in any way.